Acute brain injury can result in neuroprotection and tolerance to subsequent injury. However, the mechanisms of this endogenous neuroprotection are incompletely known. As the increase in adenosine following acute seizures is both neuroprotective and antiepileptic, adenosine may also provide neuroprotection and tolerance in ischemia. The elevation of adenosine following acute seizures is due to downregulation of adenosine kinase (ADK), the key enzyme of adenosine metabolism. Thus, the adenosine- ADK system may be a candidate as an endogenous tolerance effector. We aim to investigate how ADK is regulated in response to ischemic brain injury and how these findings can be translated into applications to prevent damage to the injured brain. The rationale for these studies is derived from the following previous findings from our lab: (1) ADK is rapidly and transiently downregulated as an acute response to both injurious seizures and transient focal cerebral ischemia. (2) Upregulation of ADK renders the brain more vulnerable to ischemic cell loss. (3) Intrastriatal implants of adenosine releasing stem cells protect the brain from subsequent ischemia. (4) Pharmacological blockade or RNAi-mediated downregulation of ADK effectively suppress seizures and seizure-induced injury. Our CENTRAL HYPOTHESIS is that the acute and transient downregulation of ADK after an insult is a general phenomenon of injury and responsible for the induction of ischemic tolerance and that augmentation and prolongation of this beneficial ADK-response is neuroprotective. We will investigate ADK expression in rodent models of ischemia. In a therapeutic approach ischemic tolerance will be induced by inhibiting ADK expression by lentiviral RNAi. The SPECIFIC AIMS of this project are: Aim 1. Study the involvement of the adenosine system in the induction of ischemic tolerance. Aim 2. Induce tolerance by therapeutic downregulation of ADK. PUBLIC HEALTH RELEVANCE: We plan to investigate novel mechanisms and strategies to prevent cell loss after stroke. Therapeutically, augmentation of the brain's endogenous neuroprotective adenosine system is expected to induce tolerance to ischemic brain injury. Our findings may be translated into effective new therapies for the prevention of brain damage after stroke.